Fluorescence is one of the most sensitive detection and imaging tools available for life science research and diagnostic assays, [1] representing the most common method for determining the presence and concentration of analytes in a wide range of applications including DNA sequencing, DNA microarrays, immunoassays, and cell imaging. The ability to detect weak signals is essential for assays requiring the detection of analytes at low concentration. To address this challenge, researchers have developed many methods to enhance fluorescence emission, thereby improving detection sensitivity. A variety of nano-patterned surfaces have been studied for the purpose of enhancing fluorescence output [2-6]. Field enhancement in these structures arises from several effects which include locally intense optical fields, reduced fluorescence lifetimes, and directional emission [7-10].
Photonic crystal (PC) surfaces comprised of a periodic surface grating coated with a high refractive index dielectric have been demonstrated for fluorescence emission enhancement applications [11] through the use of narrowband resonant modes at specific wavelengths. PC-enhanced fluorescence (PCEF) takes advantage of the resonant evanescent field that has an increased local energy density compared to the excitation light source. The intensified evanescent field strongly excites fluorophores located within an evanescent decay length of the sensor surface, resulting in enhanced emission. Previous publications have demonstrated the use of PCEF with the resonant mode spectrally overlapping the laser wavelength, to excite fluorescent dyes [12] and, that at normal incidence illumination, a PC with a resonant mode at λ=632.8 nm can produce a 60-fold magnification of cyanine-5 (Cy-5) signal compared to an ordinary glass substrate [13].
Many biological fluorescence assays, such as gene expression microarrays [14-15], have been developed using multiple fluorescent dyes within a single imaged area. In order to enhance the emission from multiple fluorescent dyes, the PC surface must be designed with resonant wavelengths coinciding with the wavelength of multiple lasers that are used to excite the target fluorophores. The PC structure intrinsically supports resonant modes in a wavelength range as wide as 200 nm [16] because each wavelength couples resonantly with the structure for a distinct angle of incidence.
Further art of interest include the published patent applications U.S. 2008/0278722 and U.S. 2006/0216204 and PCT publication WO 2008/156550.